The initial goal of our project was to identify autoantigens in type 1 diabetes. Several years ago we succeeded in cloning two novel proteins, IA-2 and IA-2 beta, from a beta cell subtraction library. These proteins were found to be enzymatically inactive members of the protein tyrosine phosphatase family, widely distributed in neuroendocrine cells throughout the body and integral transmembrane components of dense core secretory vesicles. Further studies showed they belonged to an ancient gene family going back 500 million years with homologs in Drosophila and C elegans.[unreadable] [unreadable] Examination of sera from patients with type 1 diabetes revealed that IA-2 and IA-2 beta were major autoantigens and that autoantibodies to these proteins appeared years before the development of clinical diseases. As a result, autoantibodies to IA-2 and IA-2 beta have become predictive markers for identifying individuals at high risk of developing diabetes and are being widely used to select pre-diabetics for entry into therapeutic intervention trials. Type 1 diabetes is now serving as a model for other autoimmune diseases where intensive searches for predictive autoantibodies are underway.[unreadable] [unreadable] A major focus of the project has been to determine the function of IA-2 and IA-2 beta. By knocking out these genes in mice we have shown that they are involved in the secretion of insulin and that their deletion results in a decrease in the amount of glucose-induced insulin secretion. Similar findings were observed upon knocking down IA-2 by small interfering RNA in cultured beta cells. From these and other studies we conclude that IA-2 is a modulator of both regulated and basal insulin secretion and appears to act by stabilizing dense core secretory vesicles. [unreadable] [unreadable] Studies over the last year have focused on identifying new autoantigens associated with type 1 diabetes by selective screening of proteins associated with secretory vesicles. These studies have identified two new minor autoantigens (VMP2 and NPY) thereby adding support to the hypothesis that secretory vesicle-associated proteins are particularly prone to, but not the exclusive target of, the autoimmune attack in type 1 diabetes. In addition we have developed a sensitive luminescence assay for measuring autoantibodies to IA-2. The potential clinical value of this assay is that it is robust and avoids the use of radioactivity.[unreadable] [unreadable] Natural antibodies have been an enigma because they are found in serum in the apparent absence of antigenic stimulation and are present in newborns and germ-free animals. Our earlier studies showed that polyreactive antibodies are a major component of the natural antibody repertoire, but their function has never been clearly defined. Over the last couple of year we found that polyreactive antibodies have broad antibacterial activity and with a panel of monoclonal polyreactive antibodies showed that they could bind to bacteria and acting through the classical complement pathway could inhibit the growth of Gram-positive bacteria by lysis.